PROJECT SUMMARY Aging is a key risk factor for pelvic floor disorders (PFDs), which include pelvic organ prolapse, and urinary and fecal incontinence. These common conditions interfere with the well-being of older women due to their negative impact on quality of life and associated morbidities.1 Prevalence of PFDs increase by 20% for each decade of life, reaching 50% in women age 80 and over.2 Importantly, as the U.S. population is aging, PFDs will affect an increasing proportion of women over the next decades, with a forecasted 44 million of women suffering from PFDs by 2050.2 Moreover, the substantial costs of PFDs will grow at twice the rate of the population due to the increased life expectancy in our society. Pelvic floor muscle (PFM) dysfunction is a critical defect in the progression to symptomatic PFDs.3,4 Despite this, the pathophysiology of PFDs and the cause of age-related PFM dysfunction remain poorly understood. As a result, there are currently no preventive measures and existing treatments are limited. In particular, PFM rehabilitation is less effective in older women, and surgical approaches are associated with high failure rates and adverse events.5,6 Consequently, many older women accept PFDs as an inevitable part of aging, when in fact they should demand more. It is well established that aging causes atrophy, fibrosis, and depletion of resident stem cells in limb and trunk muscles, resulting in age-related decline in muscle function.7,8,9 However, directly probing PFMs is not feasible due to their location deep within the pelvis. Current knowledge about PFMs is gleaned primarily from conventional radiological examinations that are compromised by low resolution and inability to distinguish between contractile and extracellular matrix components.10 As an alternative, human cadaveric specimens can be harnessed to begin to establish the mechanisms of age-related PFM dysfunction. The need to elucidate the pathogenesis of age-related PFDs and PFM dysfunction is clear: until basic mechanisms are established, treatments will offer marginal promise and preventive strategies will continue to be non-existent. We propose to determine functionally important age-related alterations, uncoupled from the consequences of childbirth, in the vital components of human PFMs (myofibers, extracellular matrix, stem cells). We will conduct direct examinations of cadaveric PFMs, obtained from younger and older female nulliparous donors. This project will test our overarching hypothesis that similar to other skeletal muscles, age- related atrophy, fibrosis, and diminished stem cell reservoir negatively impact PFM excursion, force production, mechanical properties, and regeneration. Our proposed cell and tissue level studies are enabled by a novel application of theoretical frameworks and advancements from other disciplines. These high-resolution analyses will facilitate the development of innovative and scientifically rationale preventive rehabilitation regiments and identification of novel therapeutic targets for mitigating age-related PFM dysfunction. Ultimately, we hope to reduce the burden of PFDs and improve the lives of millions of older women.